Acrylamide polymers are used in a wide variety of chromatographic and electrophoretic techniques and are used in capillary electrophoresis. Polyacrylamide is well suited for size fractionation of charged macro molecules such as proteins, polydeoxyribonucleic acids (DNA) and polyribonucleic acids (RNA) and when used as the matrix inside narrow bore capillaries, should greatly increase the sensitivity, efficiency, speed and resolution of electrophoretic separation and analysis of such molecules in complex mixtures.
U.S. Pat. No. 5,112,460 describes high performance micro capillary gel electrophoresis, hereby incorporated by reference. The patent indicates extremely high resolution separations of materials having different effective charges have been achieved by open tubular free-zone electrophoresis and isotachophoresis in narrow capillary tubes. In addition, bulk flow can be driven by electro osmosis to yield very sharp peaks. The patent is primarily directed toward preparing a coating material that is covalently bonded to the inner surface of the microcapillary tube prior to the preparation of the polymeric gel.
In a similar fashion, see U.S. Pat. Nos. 5,110,439 and 5,085,756, both hereby incorporated by reference.
Other references address the issue of preventing shrinkage defects during and after polymerization such as European Patent Application 272,925, published Jun. 29, 1988. Other references which are concerned with polyacrylamide gel electrophoresis are as follows: U.S. Pat. Nos. 5,141,612; 5,061,355; 5,064,769; 5,066,376; 4,997,537; 4,948,480; 4,897,306; 4,883,597; 4,874,491; 4,865,706; PCT Publication No. WO92/00795, published Jan. 23, 1992; and a paper entitled "Preparation of Polyacrylamide Gel Filled Capillaries For Capillary Electrophoresis," by Dolnik et al, published in JOURNAL OF MICROCOLUMN SEPARATIONS, Sep. 3, 1991, Vol. 3, pp. 155-159.
None of the references suggest an annealing technique of the present invention wherein subsequent to polymerization, the gel-filled capillary is cycled to an increased temperature and then cooled once and preferably several times. It is an object of the annealing technique to maintain void free gels within the capillarly tube. It is an object of the present invention to maintain the gel-filled capillaries void free particularly after the imposition of voltage during an electrophoresis separation.
The annealing makes the gel more resilient and increases its resistance to void formation during temperature changes (e.g. storage in a refrigerator) or from physical stresses, (e.g. breaking or cutting the end of capillary/gel). The annealing feature releases the stresses in the formation of the polymer.